This project investigates how human genetic variation diversifies innate immune responses to infection and cancer. Aim 1 concerns innate immunity to Epstein-Barr Virus (EBV), a cause of cancer. We found humans form two equally sized groups based on their response to EBV. Group 1 make strong Natural Killer (NK) cell and V?9V?2 T cell responses, whereas Group 2 combine a vigorous NK cell response with a poor V?9V?2 T cell response. This difference does not correlate with sex, HLA type, prior exposure to EBV or CMV, abundance of V?9V?2 T cells, or their functionality in the two groups. We propose that this even balance of responders and non-responders arises because of costs and benefits in making a V?9V?2 T cell response to EBV. Genetic precedents from our previous work are functionally distinctive haplotypes of both the killer cell immunoglobulin-like receptor (KIR) gene family, and the HLA class I gene family. We will determine genetic and functional differences that distinguish Group 1 and 2 donors, with three approaches. First, we will define the ?? T cell receptor repertoires for resting and EBV-activated ?? T cells of Group 1 and 2 donors. Second, we will define the phenotype and function of EBV-responding V?9V?2 T cells and NK cells in Group 1 and 2 individuals. Third, the ?? T cells of Group 1 and 2 individuals will be compared by high throughput RNA sequencing. Aim 2 examines NK cell and ?? T cell crosstalk in response to EBV. Our preliminary data shows that ?? T cells have an inhibitory effect on NK cell function in the context of EBV, and that this effect is distinct in Group 1 vs Group 2 donors. We propose an examination of NK cell and ?? T cell crosstalk that incorporates both NK cell educational status, and the dichotomous ?? T cell responses to EBV. Aim 3 examines interactions between KIR and HLA-A, B and C that control NK cell development and responses to infected or malignant cells that have low expression of HLA class I. These interactions involve four HLA class I epitopes and four inhibitory KIR (iKIR). The polymorphism of KIR and HLA, their independent segregation and stochastic KIR expression, mean that most humans express ligands without receptors and vice versa. During development, NK cells expressing an HLA epitope and its cognate iKIR, become educated to kill cells that lack the epitope. Consequently, educated NK cells can instigate a graft-versus-tumor (GVT) response in leukemia patients following allogeneic hematopoietic cell transplant. The GVT response clears residual tumor cells and improves patient survival. Preliminary experiments demonstrate that NK cells can be re-educated in vitro to kill malignant cells. We will determine the potential for re-education in a donor cohort representing all 12 common genetic combinations of the 4 HLA epitopes. This will give new insight on NK cell education and provide the foundation for re-educating autologous NK cells as immunotherapy for leukemia. In summary, this work will define the genetic and mechanistic parameters of three phenomena: the dichotomous V?9V?2 T cell response to EBV, V?9V?2T cell crosstalk with NK cells, and in vitro NK cell re-education.